1. Monitoring Voltage-Dependent Charge Displacement of Shaker B-IR K+ Ion Channels Using Radio Frequency Interrogation
- Author
-
Sameera Dharia and Richard D. Rabbitt
- Subjects
Anatomy and Physiology ,Patch-Clamp Techniques ,Radio Waves ,Voltage clamp ,lcsh:Medicine ,Biochemistry ,Ion Channels ,Membrane Potentials ,Biophysics Theory ,Xenopus laevis ,0302 clinical medicine ,Engineering ,Electronics Engineering ,Electricity ,Molecular Cell Biology ,Biological Systems Engineering ,Electric Impedance ,Shaker ,lcsh:Science ,Membrane potential ,Physics ,0303 health sciences ,Multidisciplinary ,Depolarization ,Animal Models ,Electrophysiology ,Signal Filtering ,Electric Field ,Membranes and Sorting ,Radio frequency ,Ion Channel Gating ,Research Article ,Biotechnology ,Biophysics ,Bioengineering ,Protein Chemistry ,Models, Biological ,03 medical and health sciences ,Model Organisms ,Electrostatics ,Electric field ,Animals ,Patch clamp ,Biology ,Ion channel ,030304 developmental biology ,lcsh:R ,Cell Membrane ,Electric Conductivity ,Proteins ,Electrophysiological Phenomena ,Signal Processing ,Oocytes ,Shaker Superfamily of Potassium Channels ,lcsh:Q ,Molecular Neuroscience ,030217 neurology & neurosurgery ,Copper ,Neuroscience - Abstract
Here we introduce a new technique that probes voltage-dependent charge displacements of excitable membrane-bound proteins using extracellularly applied radio frequency (RF, 500 kHz) electric fields. Xenopus oocytes were used as a model cell for these experiments, and were injected with cRNA encoding Shaker B-IR (ShB-IR) K(+) ion channels to express large densities of this protein in the oocyte membranes. Two-electrode voltage clamp (TEVC) was applied to command whole-cell membrane potential and to measure channel-dependent membrane currents. Simultaneously, RF electric fields were applied to perturb the membrane potential about the TEVC level and to measure voltage-dependent RF displacement currents. ShB-IR expressing oocytes showed significantly larger changes in RF displacement currents upon membrane depolarization than control oocytes. Voltage-dependent changes in RF displacement currents further increased in ShB-IR expressing oocytes after ∼120 µM Cu(2+) addition to the external bath. Cu(2+) is known to bind to the ShB-IR ion channel and inhibit Shaker K(+) conductance, indicating that changes in the RF displacement current reported here were associated with RF vibration of the Cu(2+)-linked mobile domain of the ShB-IR protein. Results demonstrate the use of extracellular RF electrodes to interrogate voltage-dependent movement of charged mobile protein domains--capabilities that might enable detection of small changes in charge distribution associated with integral membrane protein conformation and/or drug-protein interactions.
- Published
- 2011